This also refers to the same protein (locus LOC100193963) that was resembled in the BlastX search, made in October 2012, NP_001132503.1 (uncharacterized protein LOC100193963 [Zea mays], Query coverage: 99%; E value: 0.0; Max ident: 84%, and ) and where amino acid transporters conserved domains were detected.

When we aligned this amino acid sequence to the amino acid sequence of our predicted Gene IV using MultAlin we obtained the following alignment:

As it is visible, there is not a really close match that makes us state that gene IV is actually a gene. There are two clear gaps present in the alignment which is not helpful at all. Now we are looking at matches with expressed sequence tags.

2. EST Evidence

A nucleotide BLAST search against expressed sequence tags (EST database) limited for maize using the sequence for gene IV gave the following:

These particular EST results were chosen for alignment because they had the major coverage when combined. So we used both RNA sequences to make the alignment with the sequence of gene IV using MultiAlin:

We can see two big gaps (one around position 500 and the other one around position 830) and some small mismatches, which is consistent with the previous amino acid alignment. Thereby, so far we cannot tell if gene IV is actually an amino acid transporter gene because of the gaps

After some exploring, we found that there were 104 sequences that matched locus LOC100193963.This included 10 mRNA sequences and 94 est sequences. The following were found listed in UniGene under the id UGID:654651. However, the best hits are:

The first two (AFW83200.1and DAA58905.1) where submitted to NCBI on November 2012. So there are more options for making alignments with gene IV, especially with the first one because has 100% identity. Then, the MultiAlin alignment is:

Now we have a perfect match. Therefore, we can state that gene IV is an actual gene with much more confidence, although there is not RNA sequences or EST that support those two previous gaps.

3. Gene Function Evidence

The results from our RNA and est searches indicated that our Gene IV encodes a membrane amino acid transporter protein. To further investigate the possible function of Gene IV we translated our annotated nucleotide sequence to an amino acid sequence using the ExPASY translate tool and used InterPro Scan for looking conserved domains. The visual representation of the InterPro Scan results for Gene III is pasted below:

Clearly, amino acid transporter proteins coserved domains have been resembled. Therefore the function of our gene IV presumably is to encode a membrane protein involved in the transportation of amino acids into the cell.